[go: up one dir, main page]

EP1518123A1 - Electrochemical detection method and device - Google Patents

Electrochemical detection method and device

Info

Publication number
EP1518123A1
EP1518123A1 EP03740302A EP03740302A EP1518123A1 EP 1518123 A1 EP1518123 A1 EP 1518123A1 EP 03740302 A EP03740302 A EP 03740302A EP 03740302 A EP03740302 A EP 03740302A EP 1518123 A1 EP1518123 A1 EP 1518123A1
Authority
EP
European Patent Office
Prior art keywords
working electrodes
biochemical
operational amplifier
working
electrode
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
EP03740302A
Other languages
German (de)
French (fr)
Other versions
EP1518123B1 (en
Inventor
Jürgen SCHÜLEIN
Björn GRASSL
Jörg HASSMANN
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
November AG Novus Medicatus Bertling Gesellschaft fuer Molekular Medizin
Original Assignee
November AG Novus Medicatus Bertling Gesellschaft fuer Molekular Medizin
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from DE10229210A external-priority patent/DE10229210A1/en
Priority claimed from DE2002129374 external-priority patent/DE10229374C1/en
Application filed by November AG Novus Medicatus Bertling Gesellschaft fuer Molekular Medizin filed Critical November AG Novus Medicatus Bertling Gesellschaft fuer Molekular Medizin
Publication of EP1518123A1 publication Critical patent/EP1518123A1/en
Application granted granted Critical
Publication of EP1518123B1 publication Critical patent/EP1518123B1/en
Anticipated expiration legal-status Critical
Expired - Lifetime legal-status Critical Current

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
    • G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
    • G01N27/28—Electrolytic cell components
    • G01N27/30—Electrodes, e.g. test electrodes; Half-cells
    • G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
    • G01N27/3275—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction
    • G01N27/3277—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction being a redox reaction, e.g. detection by cyclic voltammetry

Definitions

  • the invention relates to a device and a method for the electrochemical detection of at least one biochemical molecule contained in a liquid from a group of predetermined biochemical molecules.
  • the invention particularly relates to a device for the detection of pathogens in a body fluid, for. B. blood.
  • Potentiostats with two or more working electrodes are used. Potentiostats with several working electrodes are also called multipotentiostats. Such multipotentiostats have a reference electrode, a counter electrode and a plurality of working electrodes. The voltage between a working electrode and the reference electrode is regulated via the voltage present between the counter electrode and the respective working electrode. A predetermined voltage curve between each of the working electrodes and the reference electrode is generated separately for each working electrode.
  • a method is known from US Pat. No. 5,830,343 in which the voltage drop across a plurality of working electrodes can be measured simultaneously using a multipotentiostat. Each working electrode is acted upon independently of the others with a special predetermined potential against the reference electrode. As a result, potentials form between the working electrodes during the measurement. This complicates the evaluation of the currents measured on the other working electrodes.
  • a method for the electrochemical detection of molecules contained in a solution is known from US Pat. No. 5,149,629, in which measurements are carried out sequentially with a plurality of working electrodes. Such a measurement is time-consuming.
  • US 4,315,753 describes a method and a device for the simultaneous determination of the concentration of second oxygen-containing gases.
  • the device has a potentiostat with two current followers which are interconnected to generate a difference signal. With the known device it is not possible to specifically detect several biochemical molecules contained in a solution.
  • US 4,655,880 discloses an apparatus for the detection of glucose. Two working electrodes are used, one of which is coated with the glucose oxidase enzyme. The other working electrode is uncoated and is used to measure the substrate. Simultaneous measurement of different biochemical molecules is not possible with the known device.
  • a biosensor is known from DE 100 15 818 AI. It is used to detect an analyte contained in a solution
  • the proposed biosensor is relatively complex to manufacture. A simultaneous detection of several biochemical molecules contained in a liquid is therefore not possible.
  • the object of the invention is to eliminate the disadvantages of the prior art.
  • a device and a method are to be specified with which a simultaneous electrochemical detection of different biochemical molecules contained in a liquid can be carried out simply, inexpensively and quickly.
  • measurement results that are as accurate as possible should be achievable.
  • a device for the electrochemical detection of a liquid contained provided with a biochemical molecule from a group of predetermined biochemical molecules is provided with a biochemical molecule from a group of predetermined biochemical molecules
  • a potentiostat for generating a predetermined voltage curve between the working electrodes and the reference electrode
  • each of the working electrodes is followed by a current-voltage converter, the current-voltage converters keeping all working electrodes at the same potential and
  • the proposed device is simple. It enables rapid detection of at least one biochemical molecule contained in a body fluid, e.g. B. a pathogen.
  • the device can be easily adapted to the biochemical molecules to be detected by selecting a suitable coating on the working electrode.
  • the type and number of the biochemical molecules contained in the "group” is given by the number of working electrodes coated with different complementary biochemical molecules.
  • the device also enables a simultaneous electrochemical detection of several different biochemical molecules contained in the liquid. Only a single potentiostat is required for the simultaneous detection of different biochemical molecules to be detected. In this way, an identical, predetermined voltage curve is applied to all working electrodes at the same time.
  • each of the working electrodes can virtually contact the circuit ground via a current follower for the individual evaluation of the signals.
  • the working electrodes are coated with biochemical molecules complementary to the biochemical molecule to be detected.
  • the working electrodes are specific for the biochemical molecules to be detected.
  • At least one specific working electrode is provided for each biochemical molecule to be detected.
  • the complementary biochemical molecules bind specifically to the biochemical molecules to be detected.
  • a “plurality of working electrodes” is understood to mean more than two working electrodes.
  • a plurality of interconnected or capacitively coupled reference electrodes are provided. This allows the speed of the measurement to continue increase.
  • a plurality of mutually connected counter electrodes can also be provided.
  • the means for measuring expediently has an analog-digital converter. Furthermore, a multiplexer can be provided, so that a quasi-simultaneous or simultaneous measurement of the currents flowing through the working electrodes is possible.
  • the current-voltage converter is a current follower having a first operational amplifier, a non-inverting input of the operational amplifier being connected to ground and the inverting input of which is connected to the output of the first operational amplifier and to the working electrode via a first resistor.
  • a capacitor can be connected in parallel with the first resistor. Noise can thus be suppressed in a simple manner and the sensitivity can thus be increased.
  • different sized first resistors can be switched on between the inverting input and the output of the first operational amplifier.
  • the current measuring range can thus be varied in a simple manner.
  • the current measuring range can be set individually for each working electrode to the optimal range for the biochemical molecule to be detected.
  • the device is universally suitable for the detection of various biochemical molecules.
  • the biochemical molecule to be detected can be a nucleic acid and the complementary biochemical molecule can be nucleic acids complementary to the nucleic acid to be detected.
  • Nucleic acids change the current profile through the corresponding working electrode. Such a change indicates that the solution contains a nucleic acid which is complementary to the nucleic acid bound to the working electrode.
  • the biochemical molecules can also be synthetic single-stranded nucleic acids or their natural and / or synthetic analogs, antigens, proteins, such as antibodies, antibody fragments, derivatives of antibodies or antibody fragments, nucleic acid-binding proteins, receptors or ligands.
  • the potentiostat has a second operational amplifier connected as a voltage follower, to the non-inverting input of which the reference electrode is connected.
  • the potentiostat can also have a third operational amplifier, to the output of which the counter electrode is connected, the inverting input of which is connected to the output of the second operational amplifier via a second resistor and via a third
  • Resistor is connected to a device for generating a selectable target voltage, and the non-inverting input of the third operational amplifier is connected to ground. Furthermore, a capacitance can be switched on between the output of the third operational amplifier and its inverting input. This causes the regulation to stabilize.
  • a method for the electrochemical detection of at least one biochemical molecule contained in a liquid from a group of predetermined biochemical molecules is provided with the following steps: a) Providing an agent for absorbing the liquid, the agent having at least one counter and a reference electrode and a plurality of working electrodes, with at least one working electrode (AE1, AE2, AE3) being provided for the detection of each biochemical molecule respective complementary molecule is coated so that the biochemical molecules can be detected simultaneously,
  • the measurement takes place almost simultaneously or simultaneously. It is expediently carried out in parallel or by means of multiplexing.
  • the voltage applied between the working electrodes and the reference electrode can be regulated with a potentiostat.
  • the proposed method is relatively easy to carry out. It is universal and also enables the simultaneous detection of a large number of different biochemical molecules in a liquid.
  • a “plurality of working electrodes” is understood to mean more than two working electrodes.
  • the predefined voltage curve can be a voltage curve that changes during the measurement.
  • the voltage curve can be specified using a programmable voltage source.
  • the electrodes can be made of conventional materials, for example suitable metals such as gold, silver, platinum or the like. be made. However, it is also possible to produce the electrodes from carbon, in particular graphite.
  • the electrodes are coated in a conventional manner, in the case of nucleic acids for example by forming covalent bonds. Reference is made to MI Pividori, et al. (2000) Electrochemical genosensor design: Immobilization of oligonucleotides onto transducer surfaces and detection methods. Biosensors and Bioelectronics 15, 291-303.
  • Fig. 1 is a schematic circuit diagram
  • FIG. 2 shows a measurement result achieved with the circuit according to FIG. 1.
  • Liquid containing molecules can be, for example, a container 1 or a field on a surface made of an insulating material, for example on a chip.
  • the container 1 has working electrodes AE1, AE2, AE3, a counter electrode GE and a reference electrode RE.
  • the electrodes are made of silver, gold, platinum or graphite, for example.
  • the working electrodes AE1, AE2, AE3 are coated with molecules that are complementary to the biochemical molecules to be detected.
  • Each of the working electrodes AE1, AE2, AE3 is connected to a measuring device AD via a current-voltage converter S1, S2, S3.
  • the current-voltage converters S1, S2, S3 each have an operational amplifier OP1, the non-inverting input (OP1 +) of which is connected to the circuit ground.
  • OP1 + the non-inverting input
  • the inverting input OP1- of the first operational amplifier OP1 is connected to the working electrode AE1, AE2, AE3 and via a first resistor R1 to the output, which in turn is connected to the measuring device AD.
  • a capacitance (not shown here) can be connected in parallel with the first resistor R1. Different sized first resistors R1 can be provided, which can alternatively be switched on. The measuring range can be changed easily.
  • the reference symbol P denotes a potentiostat, the input of which is connected to a programmable voltage source (not shown here).
  • the potentiostat P comprises a second operational amplifier OP2 connected as a voltage follower and a third operational amplifier 0P3.
  • the non-inverting input 0P2 + of the second operational amplifier OP2 is connected to the reference electrode RE.
  • the inverting input OP2- of the second operational amplifier OP2 is connected to its output and, via a second resistor, to the inverting input OP3- of the third operational amplifier OP3.
  • the non-inverting input OP3 + of the third operational amplifier is connected to the circuit ground.
  • the programmable circuit source (not shown here) is via a third resistor R3 with the inverting input OP3- of the third operational amplifier OP3 and the second resistor OP2 connected.
  • the output of the third operational amplifier OP3 is connected to the counter electrode GE.
  • a further capacitance (not shown here) can be connected between the output of the third operational amplifier OP3 and its inverting input.
  • the measuring device AD can be an analog-to-digital converter with a multiplexer. This enables a quasi-simultaneous measurement of the currents flowing through the working electrodes AE1, AE2, AE3.
  • the output of the third operational amplifier OP3 connected to the counter electrode GE is activated during operation such that no voltage is present between its inputs OP3-, OP3 +.
  • the non-inverting input 0P3 + of the third operational amplifier 0P3 is connected to the circuit ground.
  • the inverting input OP3 is also virtually at ground and thus at the same potential as the working electrodes AE1, AE2, AE3.
  • the current flowing through the third resistor R3 is equal to the current flowing through the second resistor R2.
  • the second resistor R2 is expediently selected to be the same as the third resistor R3, as a result of which the proportionality constant is fixed at the value -1.
  • the third resistor R3 can be replaced by several resistors, as a result of which several inputs, for example for modulation, are obtained.
  • FIG. 2 shows the result of measurements carried out with the circuit according to the invention.
  • uncoated working electrodes have been brought into contact with a solution containing DNA.
  • the measurement was carried out by means of differential pulse voltammetry.
  • the current difference measured at the working electrode is plotted in FIG. 2 before and after a voltage modulation.
  • the left peak shows the oxidation of guanine from DNA adsorbed on the working electrode.
  • the right peak shows the oxidation of
  • the results are plotted which have been obtained by measurement on a first working electrode AE1 and on a second working electrode AE2.
  • the present measurement only shows an unspecific detection of DNA in a solution.
  • a suitable coating of the working electrodes it is possible within the scope of the invention to detect specifically predetermined DNA or the like in a solution.
  • the number of specifically to be detected DNA sequences or the like depends on the number of working electrodes used.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Molecular Biology (AREA)
  • Physics & Mathematics (AREA)
  • General Health & Medical Sciences (AREA)
  • Immunology (AREA)
  • Electrochemistry (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • Spectroscopy & Molecular Physics (AREA)
  • General Physics & Mathematics (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Pathology (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)
  • Hybrid Cells (AREA)
  • Electric Double-Layer Capacitors Or The Like (AREA)
  • Investigating Or Analyzing Materials By The Use Of Electric Means (AREA)
  • Secondary Cells (AREA)
  • Investigating Or Analysing Biological Materials (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

The invention relates to a device for the electrochemical detection of at least one biochemical molecule-contained in a liquid-from a group of predetermined biochemical molecules, having a means ( 1 ) for taking up the liquid, said means having at least one reference electrode (RE) and at least one counterelectrode (GE) and also a multiplicity of working electrodes (AE 1 , AE 2 , AE 3 ), at least one working electrode (AE 1 , AE 2 , AE 3 ) being provided for the detection of each biochemical molecule, said working electrode being coated with a molecule that is complementary to the respective biochemical molecule, so that the biochemical molecules can be detected simultaneously, a potentiostat (P) for generating a predetermined voltage profile between the working electrodes (AE 1 , AE 2 , AE 3 ) and the reference electrode (RE), a current/voltage converter (S 1 , S 2 , S 3 ) being connected downstream of each of the working electrodes (AE 1 , AE 2 , AE 3 ), the current/voltage converters (S 1 , S 2 , S 3 ) holding all of the working electrodes (AE 1 , AE 2 , AE 3 ) at the same potential and a means (Ad) for measuring the currents flowing through the working electrodes (AE 1 , AE 2 , AE 3 ).

Description

Vorrichtung und Verfahren zum elektrochemischen NachweisDevice and method for electrochemical detection
Die Erfindung betrifft eine Vorrichtung und ein Verfahren zum elektrochemischen Nachweis zumindest eines in einer Flüssig- keit enthaltenen biochemischen Moleküls aus einer Gruppe vorgegebener biochemischer Moleküle. Die Erfindung betrifft insbesondere eine Vorrichtung zum Nachweis von Krankheitserregern in einer Körperflüssigkeit, z. B. Blut.The invention relates to a device and a method for the electrochemical detection of at least one biochemical molecule contained in a liquid from a group of predetermined biochemical molecules. The invention particularly relates to a device for the detection of pathogens in a body fluid, for. B. blood.
Zur Messung elektrochemischer Potenziale werden nach demFor the measurement of electrochemical potentials after the
Stand der Technik Potentiostaten mit zwei oder mehreren Arbeitselektroden verwendet. Potentiostaten mit mehreren Arbeitselektroden werden auch als Multipotentiostaten bezeichnet. Solche Multipotentiostaten weisen eine Referenzelektro- de, eine Gegenelektrode und mehrere Arbeitselektroden auf. Die Spannung zwischen einer Arbeitselektrode und der Referenzelektrode wird über die zwischen der Gegenelektrode und der jeweiligen Arbeitselektrode anliegende Spannung geregelt. Ein vorgegebener Spannungsverlauf zwischen jeder der Arbeit- selektroden und der Referenzelektrode wird für jede Arbeitselektrode separat erzeugt.State of the art potentiostats with two or more working electrodes are used. Potentiostats with several working electrodes are also called multipotentiostats. Such multipotentiostats have a reference electrode, a counter electrode and a plurality of working electrodes. The voltage between a working electrode and the reference electrode is regulated via the voltage present between the counter electrode and the respective working electrode. A predetermined voltage curve between each of the working electrodes and the reference electrode is generated separately for each working electrode.
Aus der US 5,830,343 ist ein Verfahren bekannt, bei dem mittels eines Multipotentiostaten simultan die über eine Viel- zahl von Arbeitselektroden abfallende Spannung gemessen werden kann. Dabei wird jede Arbeitselektrode unabhängig von den anderen mit einem besonderen vorgegebenen Potenzial gegen die Referenzelektrode beaufschlagt. Infolgedessen bilden sich während der Messung zwischen den Arbeitselektroden Potenziale aus. Das macht die Auswertung der an den übrigen Arbeitselektroden gemessenen Ströme kompliziert. Aus der US 5,149,629 ist ein Verfahren zur elektrochemischen Detektion von in einer Lösung enthaltenen Molekülen bekannt, bei dem sequenziell mit mehreren Arbeitselektroden gemessen wird. Die Durchführung einer solchen Messung ist zeitaufwän- dig.A method is known from US Pat. No. 5,830,343 in which the voltage drop across a plurality of working electrodes can be measured simultaneously using a multipotentiostat. Each working electrode is acted upon independently of the others with a special predetermined potential against the reference electrode. As a result, potentials form between the working electrodes during the measurement. This complicates the evaluation of the currents measured on the other working electrodes. A method for the electrochemical detection of molecules contained in a solution is known from US Pat. No. 5,149,629, in which measurements are carried out sequentially with a plurality of working electrodes. Such a measurement is time-consuming.
Die US 4,315,753 beschreibt ein Verfahren und eine Vorrichtung zur gleichzeitigen Ermittlung der Konzentration zweiter Sauerstoffhaltiger Gase. Die Vorrichtung weist einen Poten- tiostaten mit zwei Stromfolgern auf, die zur Erzeugung eines Differenzsignals miteinander verschalten sind. Mit der bekannten Vorrichtung ist es nicht möglich, mehrere in einer Lösung enthaltene biochemische Moleküle spezifisch nachzuweisen.US 4,315,753 describes a method and a device for the simultaneous determination of the concentration of second oxygen-containing gases. The device has a potentiostat with two current followers which are interconnected to generate a difference signal. With the known device it is not possible to specifically detect several biochemical molecules contained in a solution.
Die US 4,655,880 offenbart eine Vorrichtung zum Nachweis von Glucose. Dabei werden zwei Arbeitselektroden verwendet, von denen eine mit dem Glucose-Oxidase-Enzym beschichtet ist. Die andere Arbeitselektrode ist unbeschichtet und dient der Mes- sung des Untergrunds. Eine simultane Messung unterschiedlicher biochemischer Moleküle ist mit der bekannten Vorrichtung nicht möglich.US 4,655,880 discloses an apparatus for the detection of glucose. Two working electrodes are used, one of which is coated with the glucose oxidase enzyme. The other working electrode is uncoated and is used to measure the substrate. Simultaneous measurement of different biochemical molecules is not possible with the known device.
Aus Paeschke, Manfred et al . : Voltammetric Multichannel Meas- urements Using Silicon Fabricated Microelectrode Arrays; in: Electroanalysis 1996, 8, Nr. 10; Seiten 891 bis 898 ist ein voltammetrisches Verfahren unter Verwendung eines Vielkanal - potentiostaten beschrieben. Der beschriebene Vielkanalpoten- tiostat ist aufwändig herzustellen. Angesehen davon, ergeben sich in der Praxis bei der Messung häufig Stabilitätsprobleme. Ein spezifischer Nachweis von in einer Lösung enthaltenen biochemischen Molekülen ist damit nur eingeschränkt möglich. Die DE 41 36 779 AI beschreibt eine Vorrichtung zum simultanen Nachweis verschiedener Gaskomponenten. Die Vorrichtung umfasst verschiedene Arbeitselektroden, eine gemeinsame Gegenelektrode sowie eine gemeinsame Referenzelektrode. Mit der Vorrichtung kann das Potenzial jeder Arbeitselektrode separat geregelt werden. Die entsprechende Regelungsschaltung ist kompliziert und störanfällig.From Paeschke, Manfred et al. : Voltammetric Multichannel Measurements Using Silicon Fabricated Microelectrode Arrays; in: Electroanalysis 1996, 8, No. 10; Pages 891 to 898 describe a voltammetric method using a multi-channel potentiostat. The multi-channel potentiostat described is complex to manufacture. Apart from this, stability problems often arise in practice during the measurement. Specific detection of biochemical molecules contained in a solution is therefore only possible to a limited extent. DE 41 36 779 AI describes a device for the simultaneous detection of different gas components. The device comprises various working electrodes, a common counter electrode and a common reference electrode. With the device, the potential of each working electrode can be regulated separately. The corresponding control circuit is complicated and prone to failure.
Aus der DE 100 15 818 AI ist ein Biosensor bekannt. Zum Nach- weis eines in einer Lösung enthaltenen Analyten wird dessenA biosensor is known from DE 100 15 818 AI. It is used to detect an analyte contained in a solution
Oxidations- und Reduktionspotential an jeweils einer Elektrode gemessen. Es sind also zwei Elektroden pro nachzuweisendem Analyt notwendig. Der vorgeschlagene Biosensor ist relativ aufwändig herzustellen. Ein gleichzeitiger Nachweis mehrerer in einer Flüssigkeit enthaltener biochemischer Moleküle ist damit nicht möglich.Oxidation and reduction potential measured on one electrode each. Two electrodes are therefore required per analyte to be detected. The proposed biosensor is relatively complex to manufacture. A simultaneous detection of several biochemical molecules contained in a liquid is therefore not possible.
Aufgabe der Erfindung ist es, die Nachteile nach dem Stand der Technik zu beseitigen. Es sollen insbesondere eine Vor- richtung und ein Verfahren angegeben werden, mit denen ein simultaner elektrochemischer Nachweis von in einer Flüssigkeit enthaltenen unterschiedlichen biochemischen Molekülen einfach, kostengünstig und schnell durchführbar ist. Nach einem weiteren Ziel der Erfindung sollen möglichst genaue Mes- sergebnisse erzielbar sein.The object of the invention is to eliminate the disadvantages of the prior art. In particular, a device and a method are to be specified with which a simultaneous electrochemical detection of different biochemical molecules contained in a liquid can be carried out simply, inexpensively and quickly. According to a further object of the invention, measurement results that are as accurate as possible should be achievable.
Diese Aufgabe wird durch die Merkmale der Ansprüche 1 und 12 gelöst. Zweckmäßige Ausgestaltungen ergeben sich aus den Merkmalen der Ansprüche 2 bis 11, 13 und 14.This object is solved by the features of claims 1 and 12. Appropriate configurations result from the features of claims 2 to 11, 13 and 14.
Nach Maßgabe der Erfindung ist eine Vorrichtung zum elektrochemischen Nachweis eines in einer Flüssigkeit enthaltenen biochemischen Moleküls aus einer Gruppe vorgegebener biochemischer Moleküle vorgesehen mitAccording to the invention, a device for the electrochemical detection of a liquid contained provided with a biochemical molecule from a group of predetermined biochemical molecules
einem mindestens eine Referenz- und mindestens eine Gegene- lektrode sowie eine Vielzahl an Arbeitselektroden aufweisenden Mittel zur Aufnahme der Flüssigkeit, wobei zum Nachweis jedes biochemischen Moleküls zumindest eine Arbeitselektrode (AEl, AE2 , AE3) vorgesehen ist, die mit einem zum jeweiligen biochemischen Molekül komplementären Molekül beschichtet ist, so dass die biochemischen Moleküle simultan nachweisbar sind,at least one reference electrode and at least one counter electrode and a plurality of working electrodes having means for absorbing the liquid, with at least one working electrode (AE1, AE2, AE3) being provided for the detection of each biochemical molecule, which electrode is complementary to the respective biochemical molecule Molecule is coated so that the biochemical molecules can be detected simultaneously,
einem Potentiostaten zur Erzeugung eines vorgegebenen Spannungsverlaufs zwischen den Arbeitselektroden und der Referenzelektrode ,a potentiostat for generating a predetermined voltage curve between the working electrodes and the reference electrode,
wobei jeder der Arbeitselektroden ein Strom-Spannungskonverter nachgeschaltet ist, wobei die Strom-Spannungskonverter sämtliche Arbeitselektroden auf demselben Potenzial halten undwherein each of the working electrodes is followed by a current-voltage converter, the current-voltage converters keeping all working electrodes at the same potential and
einem Mittel zum Messen der durch die Arbeitselektroden fließenden Ströme.a means for measuring the currents flowing through the working electrodes.
Die vorgeschlagene Vorrichtung ist einfach aufgebaut. Sie er- möglicht einen schnellen Nachweis zumindest eines in einer Körperflüssigkeit enthaltenen biochemischen Moleküls, z. B. eines Krankheitserregers. Die Vorrichtung lässt sich einfach durch die Wahl einer geeigneten Beschichtung der Arbeitselektrode an die nachzuweisenden biochemischen Moleküle anpassen. Die Art und die Anzahl der in der "Gruppe" enthaltenen biochemischen Moleküle ist durch die Anzahl der mit unterschiedlichen komplementären biochemischen Molekülen beschichteten Arbeitselektroden gegeben. Die Vorrichtung ermöglicht auch einen simultanen elektrochemischen Nachweis mehrerer unterschiedlicher in der Flüssigkeit enthaltener biochemischer Moleküle. Zum simultanen Nachweis unterschiedlicher nachzuweisender biochemischer Moleküle ist lediglich ein einziger Po- tentiostat erforderlich. Damit wird an sämtliche Arbeitselektroden gleichzeitig ein identischer vorgegebener Spannungsverlauf angelegt. Indem sämtliche Arbeitselektroden auf demselben Potenzial gehalten werden, ist es möglich, die durch die Arbeitselektroden fließenden Ströme parallel zu messen. Dazu kann jede der Arbeitselektroden über einen Stromfolger zur individuellen Auswertung der Signale virtuell an der Schaltungsmasse anliegen. Zum spezifischen Nachweis der in der Flüssigkeit enthaltenen biochemischen Moleküle sind die Arbeitselektroden mit zum nachzuweisenden biochemischen Mole- kül komplementären biochemischen Molekülen beschichtet. Die Arbeitselektroden sind spezifisch für die nachzuweisenden biochemischen Molekülen. Für jedes nachzuweisende biochemische Molekül ist zumindest eine spezifische Arbeitselektrode vorgesehen. Die komplementären biochemischen Moleküle binden spezifisch an den nachzuweisenden biochemischen Moleküle. Infolge der Ausbildung einer aus dem nachzuweisenden biochemischen Molekül und dem komplementären biochemischen Molekül gebildeten Verbindung ändert sich das elektrochemische Signal der Arbeitselektrode.The proposed device is simple. It enables rapid detection of at least one biochemical molecule contained in a body fluid, e.g. B. a pathogen. The device can be easily adapted to the biochemical molecules to be detected by selecting a suitable coating on the working electrode. The type and number of the biochemical molecules contained in the "group" is given by the number of working electrodes coated with different complementary biochemical molecules. The device also enables a simultaneous electrochemical detection of several different biochemical molecules contained in the liquid. Only a single potentiostat is required for the simultaneous detection of different biochemical molecules to be detected. In this way, an identical, predetermined voltage curve is applied to all working electrodes at the same time. By keeping all working electrodes at the same potential, it is possible to measure the currents flowing through the working electrodes in parallel. For this purpose, each of the working electrodes can virtually contact the circuit ground via a current follower for the individual evaluation of the signals. For specific detection of the biochemical molecules contained in the liquid, the working electrodes are coated with biochemical molecules complementary to the biochemical molecule to be detected. The working electrodes are specific for the biochemical molecules to be detected. At least one specific working electrode is provided for each biochemical molecule to be detected. The complementary biochemical molecules bind specifically to the biochemical molecules to be detected. As a result of the formation of a compound formed from the biochemical molecule to be detected and the complementary biochemical molecule, the electrochemical signal of the working electrode changes.
Unter einer "Vielzahl von Arbeitselektroden" werden im Sinne der vorliegenden Erfindung mehr als zwei Arbeitselektroden verstanden.For the purposes of the present invention, a “plurality of working electrodes” is understood to mean more than two working electrodes.
Nach einer vorteilhaften Ausgestaltung sind mehrere miteinander verbundene oder kapazitiv gekoppelte Referenzelektroden vorgesehen. Damit kann die Geschwindigkeit der Messung weiter erhöht werden. In diesem Zusammenhang können auch mehrere miteinander verbundene Gegenelektroden vorgesehen sein.According to an advantageous embodiment, a plurality of interconnected or capacitively coupled reference electrodes are provided. This allows the speed of the measurement to continue increase. In this context, a plurality of mutually connected counter electrodes can also be provided.
Zweckmäßigerweise weist das Mittel zum Messen einen Analog- Digital-Wandler auf. Ferner kann ein Multiplexer vorgesehen sein, so dass eine quasi zeitgleiche bzw. simultane Messung der durch die Arbeitselektroden fließenden Ströme möglich ist.The means for measuring expediently has an analog-digital converter. Furthermore, a multiplexer can be provided, so that a quasi-simultaneous or simultaneous measurement of the currents flowing through the working electrodes is possible.
Nach einer weiteren Ausgestaltung ist der Strom-Spannungskonverter ein einen ersten Operationsverstärker aufweisender Stromfolger, wobei ein nichtinvertierender Eingang des Operationsverstärkers an der Masse anliegt und dessen invertierender Eingang über einen ersten Widerstand mit dem Ausgang des ersten Operationsverstärkers und mit der Arbeitselektrode verbunden ist. Parallel zum ersten Widerstand kann eine Kapazität geschaltet sein. Damit kann auf einfache Weise ein Rauschen unterdrücktund somit die Sensitivität gesteigert werden. Zur Einstellung des Strommessbereichs können unterschiedlich große erste Widerstände zwischen dem invertierenden Eingang und dem Ausgang des ersten Operationsverstärkers einschaltbar sein. Damit kann in einfacher Weise der Strommessbereich variiert werden. Der Strommessbereich kann für jede Arbeits- elektrode individuell auf den für das nachzuweisende biochemische Molekül optimalen Bereich eingestellt werden. Die Vorrichtung ist universell für den Nachweis unterschiedlichster biochemischer Moleküle geeignet.According to a further embodiment, the current-voltage converter is a current follower having a first operational amplifier, a non-inverting input of the operational amplifier being connected to ground and the inverting input of which is connected to the output of the first operational amplifier and to the working electrode via a first resistor. A capacitor can be connected in parallel with the first resistor. Noise can thus be suppressed in a simple manner and the sensitivity can thus be increased. To set the current measuring range, different sized first resistors can be switched on between the inverting input and the output of the first operational amplifier. The current measuring range can thus be varied in a simple manner. The current measuring range can be set individually for each working electrode to the optimal range for the biochemical molecule to be detected. The device is universally suitable for the detection of various biochemical molecules.
Beim nachzuweisenden biochemischen Molekül kann es sich um eine Nukleinsäure und beim komplementären biochemischen Molekül um zur nachzuweisenden Nukleinsäure komplementäre Nukleinsäuren handeln. Im Falle einer Hybridisierung solcher Nukleinsäuren ändert sich der Stromverlauf durch die entsprechende Arbeitselektrode. Eine solche Änderung zeigt an, dass in der Lösung eine Nukleinsäure enthalten ist, welche zur an der Arbeitselektrode gebundenen Nukleinsäure komplementär ist. Ein solcher Nachweis ist hoch sensitiv und äußerst spezifisch. Bei den biochemischen Molekülen kann es sich auch um synthetische einzelsträngige Nukleinsäuren oder deren natürliche und/oder synthetische Analoga, Antigene, Proteine, wie Antikörper, Antikörperfragmente, Derivate von Antikörpern oder Antikörperfragmenten, Nukleinsäure-bindende Proteine, Rezeptoren oder Liganden handeln.The biochemical molecule to be detected can be a nucleic acid and the complementary biochemical molecule can be nucleic acids complementary to the nucleic acid to be detected. In the case of hybridization such Nucleic acids change the current profile through the corresponding working electrode. Such a change indicates that the solution contains a nucleic acid which is complementary to the nucleic acid bound to the working electrode. Such evidence is highly sensitive and extremely specific. The biochemical molecules can also be synthetic single-stranded nucleic acids or their natural and / or synthetic analogs, antigens, proteins, such as antibodies, antibody fragments, derivatives of antibodies or antibody fragments, nucleic acid-binding proteins, receptors or ligands.
In weiterer Ausgestaltung weist der Potentiostat einen als Spannungsfolger geschalteten zweiten Operationsverstärker auf, an dessen nicht invertierendem Eingang die Referenzelektrode angeschlossen ist. Der Potentiostat kann ferner einen dritten Operationsverstärker aufweisen, an dessen Ausgang die Gegenelektrode angeschlossen ist, dessen invertierender Eingang über einen zweiten Widerstand mit dem Ausgang des zwei- ten Operationsverstärkers verbunden und über einen drittenIn a further embodiment, the potentiostat has a second operational amplifier connected as a voltage follower, to the non-inverting input of which the reference electrode is connected. The potentiostat can also have a third operational amplifier, to the output of which the counter electrode is connected, the inverting input of which is connected to the output of the second operational amplifier via a second resistor and via a third
Widerstand mit einer Einrichtung zur Erzeugung einer wählbaren Sollspannung angeschlossen ist, und wobei der nichtinver- tierende Eingang des dritten Operationsverstärkers an der Masse anliegt. Des Weiteren kann zwischen dem Ausgang des dritten Operationsverstärkers und dessen invertierenden Eingang eine Kapazität eingeschaltet sein. Das bewirkt eine Stabilisierung der Regelung.Resistor is connected to a device for generating a selectable target voltage, and the non-inverting input of the third operational amplifier is connected to ground. Furthermore, a capacitance can be switched on between the output of the third operational amplifier and its inverting input. This causes the regulation to stabilize.
Nach weiterer Maßgabe der Erfindung ist ein Verfahren zum elektrochemischen Nachweis zumindest eines in einer Flüssigkeit enthaltenen biochemischen Moleküls aus einer Gruppe vorgegebener biochemischer Moleküle mit folgenden Schritten vorgesehen: a) Bereitstellen eines Mittels zur Aufnahme der Flüssigkeit, wobei das Mittel mindestens eine Gegen- und eine Referenzelektrode sowie eine Vielzahl von Arbeitselektroden aufweist, wobei zum Nachweis jedes biochemischen Moleküls zumindest eine Arbeitselektrode (AEl, AE2 , AE3) vorgesehen ist, die mit einem zum jeweiligen biochemischen Molekül komplementären Molekül beschichtet ist, so dass die biochemischen Moleküle simultan nachweisbar sind,According to a further aspect of the invention, a method for the electrochemical detection of at least one biochemical molecule contained in a liquid from a group of predetermined biochemical molecules is provided with the following steps: a) Providing an agent for absorbing the liquid, the agent having at least one counter and a reference electrode and a plurality of working electrodes, with at least one working electrode (AE1, AE2, AE3) being provided for the detection of each biochemical molecule respective complementary molecule is coated so that the biochemical molecules can be detected simultaneously,
b) Inkontaktbringen der Flüssigkeit mit den Arbeits-, Gegen- und Referenzelektroden,b) bringing the liquid into contact with the working, counter and reference electrodes,
c) gleichzeitiges Anlegen eines vorgegebenen Spannungsver- laufs zwischen den Arbeitselektroden und der Referenzelektrode undc) simultaneous application of a predetermined voltage profile between the working electrodes and the reference electrode and
d) Messen der durch die Arbeitselektroden fließenden Ströme, wobei während der Messung sämtliche Arbeitselektro- den auf demselben Potenzial gehalten werden.d) measuring the currents flowing through the working electrodes, all working electrodes being kept at the same potential during the measurement.
Die Messung erfolgt quasi zeitgleich bzw. simultan. Sie wird zweckmäßigerweise parallel oder mittels Multiplexen durchgeführt. Dabei kann die zwischen den Arbeitselektroden und der Referenzelektrode anliegende Spannung mit einem Potentiostaten geregelt werden. Das vorgeschlagene Verfahren ist relativ einfach durchführbar. Es ist universell und ermöglicht auch den simultanen Nachweis einer Vielzahl unterschiedlicher biochemischer Moleküle in einer Flüssigkeit.The measurement takes place almost simultaneously or simultaneously. It is expediently carried out in parallel or by means of multiplexing. The voltage applied between the working electrodes and the reference electrode can be regulated with a potentiostat. The proposed method is relatively easy to carry out. It is universal and also enables the simultaneous detection of a large number of different biochemical molecules in a liquid.
Unter einer "Vielzahl von Arbeitselektroden" werden im Sinne der vorliegenden Erfindung mehr als zwei Arbeitselektroden verstanden. Bei dem vorgegebenen Spannungsverlauf kann es sich um einen sich während der Messung veränderlichen Spannungsverlauf handeln. Der Spannungsverlauf kann mittels einer programmierbaren Spannungsquelle vorgegeben werden.For the purposes of the present invention, a “plurality of working electrodes” is understood to mean more than two working electrodes. The predefined voltage curve can be a voltage curve that changes during the measurement. The voltage curve can be specified using a programmable voltage source.
Die Elektroden können aus herkömmlichen Materialien, beispielsweise geeigneten Metallen wie Gold, Silber, Platin oder dgl . hergestellt sein. Es ist aber auch möglich, die Elektroden aus Kohlenstoff, insbesondere Grafit, herzustellen. Die Beschichtung der Elektroden erfolgt in herkömmlicher Weise, im Falle von Nukleinsäuren beispielsweise durch Ausbildung von kovalenten Bindungen. Es wird verwiesen auf MI Pividori , et al . (2000) Electrochemical genosensor design : Immobilisa - tion of oligonucleotides onto transducer surfaces and detec- tion methods . Biosensors and Bioelectronics 15, 291 -303 .The electrodes can be made of conventional materials, for example suitable metals such as gold, silver, platinum or the like. be made. However, it is also possible to produce the electrodes from carbon, in particular graphite. The electrodes are coated in a conventional manner, in the case of nucleic acids for example by forming covalent bonds. Reference is made to MI Pividori, et al. (2000) Electrochemical genosensor design: Immobilization of oligonucleotides onto transducer surfaces and detection methods. Biosensors and Bioelectronics 15, 291-303.
Ein Ausführungsbeispiel der Erfindung wird anhand der Zeichnung näher erläutert. Es zeigen:An embodiment of the invention is explained in more detail with reference to the drawing. Show it:
Fig. 1 ein schematisches Schaltbild undFig. 1 is a schematic circuit diagram and
Fig. 2 ein mit der Schaltung gemäß Fig. 1 erzieltes Messergebnis .FIG. 2 shows a measurement result achieved with the circuit according to FIG. 1.
Ein Mittel zur Aufnahme der die nachzuweisenden biochemischenA means of recording the biochemical to be detected
Moleküle enthaltenden Flüssigkeit kann z.B. ein Behälter 1 oder ein Feld auf einer aus einem isolierenden Material hergestellten Fläche, z.B. auf einem Chip, sein. Der Behälter 1 weist Arbeitselektroden AEl, AE2 , AE3 , eine Gegenelektrode GE sowie eine Referenzelektrode RE auf. Die Elektroden sind z.B. aus Silber, Gold, Platin oder Grafit hergestellt. Die Arbeitselektroden AEl, AE2 , AE3 sind mit zu den nachzuweisenden biochemischen Molekülen komplementären Moleküle beschichtet. Jede der Arbeitselektroden AEl, AE2 , AE3 ist über eine Strom- Spannungskonverter Sl, S2, S3 mit einer Messvorrichtung AD verbunden.Liquid containing molecules can be, for example, a container 1 or a field on a surface made of an insulating material, for example on a chip. The container 1 has working electrodes AE1, AE2, AE3, a counter electrode GE and a reference electrode RE. The electrodes are made of silver, gold, platinum or graphite, for example. The working electrodes AE1, AE2, AE3 are coated with molecules that are complementary to the biochemical molecules to be detected. Each of the working electrodes AE1, AE2, AE3 is connected to a measuring device AD via a current-voltage converter S1, S2, S3.
Die Strom-Spannungskonverter Sl, S2 , S3 weisen jeweils einen Operationsverstärker OP1 auf, dessen nichtinvertierender Eingang (OP1+) an Schaltungsmasse anliegt. Infolgedessen werden sämtliche Arbeitselektroden AEl, AE2 , AE3 auf demselben Potenzial gehalten. Der invertierende Eingang OP1- des ersten Operationsverstärkers OP1 ist mit der Arbeitselektrode AEl, AE2 , AE3 und über einen ersten Widerstand Rl mit dem Ausgang verbunden, der wiederum mit der Messvorrichtung AD in Verbindung steht. Zur Rauschunterdrückung kann parallel zum ersten Widerstand Rl eine (hier nicht gezeigte) Kapazität geschaltet sein. Es können unterschiedlich große erste Widerstände Rl vorgesehen sein, welche alternativ einschaltbar sind. So kann auf einfache Weise der Messbereich geändert werden.The current-voltage converters S1, S2, S3 each have an operational amplifier OP1, the non-inverting input (OP1 +) of which is connected to the circuit ground. As a result, all working electrodes AE1, AE2, AE3 are kept at the same potential. The inverting input OP1- of the first operational amplifier OP1 is connected to the working electrode AE1, AE2, AE3 and via a first resistor R1 to the output, which in turn is connected to the measuring device AD. To suppress noise, a capacitance (not shown here) can be connected in parallel with the first resistor R1. Different sized first resistors R1 can be provided, which can alternatively be switched on. The measuring range can be changed easily.
Mit dem Bezugszeichen P ist ein Potentiostat bezeichnet, des- sen Eingang mit einer (hier nicht gezeigten) programmierbaren Spannungsquelle verbunden ist. Der Potentiostat P umfasst einen als Spannungsfolger geschalteten zweiten Operationsverstärker OP2 und einen dritten Operationsverstärker 0P3. Der nichtinvertierende Eingang 0P2+ des zweiten Operationsver- stärkers OP2 ist an die Referenzelektrode RE angeschlossen.The reference symbol P denotes a potentiostat, the input of which is connected to a programmable voltage source (not shown here). The potentiostat P comprises a second operational amplifier OP2 connected as a voltage follower and a third operational amplifier 0P3. The non-inverting input 0P2 + of the second operational amplifier OP2 is connected to the reference electrode RE.
Der invertierende Eingang OP2- des zweiten Operationsverstärkers OP2 ist mit dessen Ausgang und über einen zweiten Widerstand mit dem invertierenden Eingang OP3 - des dritten Operationsverstärkers OP3 verbunden. Der nichtinvertierende Ein- gang OP3+ des dritten Operationsverstärkers liegt an Schaltungsmasse. Die (hier nicht gezeigte) programmierbare Schaltungsquelle ist über einen dritten Widerstand R3 mit dem invertierenden Eingang OP3- des dritten Operationsverstärkers OP3 sowie dem zweiten Widerstand OP2 verbunden. Der Ausgang des dritten Operationsverstärkers OP3 ist mit der Gegenelektrode GE verbunden. Zwischen dem Ausgang des dritten Operationsverstärkers OP3 und dessen invertierenden Eingang kann ei- ne (hier nicht gezeigte) weitere Kapazität eingeschaltet sein.The inverting input OP2- of the second operational amplifier OP2 is connected to its output and, via a second resistor, to the inverting input OP3- of the third operational amplifier OP3. The non-inverting input OP3 + of the third operational amplifier is connected to the circuit ground. The programmable circuit source (not shown here) is via a third resistor R3 with the inverting input OP3- of the third operational amplifier OP3 and the second resistor OP2 connected. The output of the third operational amplifier OP3 is connected to the counter electrode GE. A further capacitance (not shown here) can be connected between the output of the third operational amplifier OP3 and its inverting input.
Bei der Messvorrichtung AD kann es sich um einen Analog-Digital -Wandler mit Multiplexer handeln. Das ermöglicht eine quasi zeitgleiche Messung der durch die Arbeitselektroden AEl, AE2, AE3 fließenden Ströme.The measuring device AD can be an analog-to-digital converter with a multiplexer. This enables a quasi-simultaneous measurement of the currents flowing through the working electrodes AE1, AE2, AE3.
Indem die Referenzelektrode RE an den nichtinvertierenden Eingang OP2+ des zweiten Operationsverstärkers OP2 ange- schlössen ist, erhält man einen Spannungsfolger mit einer sehr hohen Eingangsimpedanz. Ein durch die Referenzelektrode RE fließender Elektrolysestrom wird damit wirkungsvoll unterbunden. Infolgedessen wird eine besonders genaue Messung erreicht .By connecting the reference electrode RE to the non-inverting input OP2 + of the second operational amplifier OP2, a voltage follower with a very high input impedance is obtained. An electrolysis current flowing through the reference electrode RE is thus effectively prevented. As a result, a particularly accurate measurement is achieved.
Der mit der Gegenelektrode GE verbundene Ausgang des dritten Operationsverstärkers OP3 wird im Betrieb so angesteuert, dass zwischen dessen Eingängen OP3-, OP3+ keine Spannung anliegt. Der nichtinvertierende Eingang 0P3+ des dritten Opera- tionsverstarkers 0P3 liegt an Schaltungsmasse an. Infolgedessen liegt auch der invertierende Eingang OP3- virtuell auf Masse und damit auf demselben Potenzial wie die Arbeitselektroden AEl, AE2 , AE3. Bei geeigneter Regelung ist der durch den dritten Widerstand R3 fließende Strom gleich dem durch den zweiten Widerstand R2 fließenden Strom. Da die Spannung über dem zweiten Widerstand R2 dem Betrag nach gleich der Spannung zwischen der Referenzelektrode RE und den Arbeitselektroden AEl, AE2 , AE3 ist, kann das Potenzial der Arbeitse- lektroden AEl, AE2 , AE3 gegen die Referenzelektrode RE durch eine proportionale Spannung am Eingang U des Potentiostaten P vorgegeben werden. In der Praxis wird zweckmäßigerweise der zweite Widerstand R2 gleich dem dritten Widerstand R3 gewählt, wodurch die Proportionalitätskonstante auf den Wert -1 festgelegt wird. Alternativ kann in diesem Fall eines addierenden Potentiostaten P der dritte Widerstand R3 durch mehrere Widerstände ersetzt werden, wodurch mehrere Eingänge, z.B. zur Modulation, erhalten werden.The output of the third operational amplifier OP3 connected to the counter electrode GE is activated during operation such that no voltage is present between its inputs OP3-, OP3 +. The non-inverting input 0P3 + of the third operational amplifier 0P3 is connected to the circuit ground. As a result, the inverting input OP3 is also virtually at ground and thus at the same potential as the working electrodes AE1, AE2, AE3. With suitable regulation, the current flowing through the third resistor R3 is equal to the current flowing through the second resistor R2. Since the voltage across the second resistor R2 is equal in magnitude to the voltage between the reference electrode RE and the working electrodes AE1, AE2, AE3, the potential of the working electrodes AE1, AE2, AE3 against the reference electrode RE by a proportional voltage at the input U of the potentiostat P can be specified. In practice, the second resistor R2 is expediently selected to be the same as the third resistor R3, as a result of which the proportionality constant is fixed at the value -1. Alternatively, in this case of an adding potentiostat P, the third resistor R3 can be replaced by several resistors, as a result of which several inputs, for example for modulation, are obtained.
Nach einer Variation an der Schaltung ist es möglich, die Strom-Spannungskonverter Sl, S2 , S3 im Frequenzgang zu beschränken. Dadurch kann das Gesamtrauschen vermindert werden. Eine solche Beschränkung im Frequenzgang kann durch Kondensa- toren erreicht werden, die jeweils parallel zum ersten Widerstand Rl geschaltet werden. Zur Vergrößerung des Strommessbereichs kann es von Vorteil sein, die ersten Widerstände Rl , ggf. mit dazu parallel geschalteten Kondensatoren, mittels Relais oder analoger elektronischer Schalter oder einer Kom- bination der beiden schaltbar auszuführen.After a variation on the circuit, it is possible to limit the current-voltage converters S1, S2, S3 in the frequency response. This can reduce the overall noise. Such a limitation in the frequency response can be achieved by capacitors, which are each connected in parallel to the first resistor R1. To increase the current measuring range, it can be advantageous to design the first resistors R1, possibly with capacitors connected in parallel, by means of relays or analog electronic switches or a combination of the two.
Fig. 2 zeigt das Ergebnis von mit der erfindungsgemäßen Schaltung durchgeführten Messungen. Dazu sind unbeschichtete Arbeitselektroden mit einer DNA enthaltenden Lösung in Kon- takt gebracht worden. Die Messung ist erfolgt mittels Diffe- renzialpulsvoltammetrie . Aufgetragen in Fig. 2 ist über der Spannung die an der Arbeitselektrode gemessene Stromdifferenz jeweils vor und nach einer Spannungsmodulation. Der linke Pe- ak zeigt die Oxidation von Guanin von an der Arbeitselektrode adsorbierter DNA. Der rechte Peak zeigt die Oxidation von2 shows the result of measurements carried out with the circuit according to the invention. For this purpose, uncoated working electrodes have been brought into contact with a solution containing DNA. The measurement was carried out by means of differential pulse voltammetry. The current difference measured at the working electrode is plotted in FIG. 2 before and after a voltage modulation. The left peak shows the oxidation of guanine from DNA adsorbed on the working electrode. The right peak shows the oxidation of
Adenin. Es sind die Ergebnisse aufgetragen welche durch Messung an einer ersten Arbeitselektrode AEl und an einer zweiten Arbeitselektrode AE2 gewonnen worden sind. Die vorliegende Messung zeigt lediglich einen unspezifischen Nachweis von DNA in einer Lösung. Bei einer geeigneten Beschichtung der Arbeitselektroden ist es im Rahmen der Erfindung möglich, spezifisch vorgegebene DNA oder dgl. in einer Lösung nachzuweisen. Die Anzahl der spezifisch nachzuweisenden DNA-Sequenzen oder dgl. hängt ab von der Anzahl der verwendeten Arbeitselektroden. Adenine. The results are plotted which have been obtained by measurement on a first working electrode AE1 and on a second working electrode AE2. The present measurement only shows an unspecific detection of DNA in a solution. With a suitable coating of the working electrodes, it is possible within the scope of the invention to detect specifically predetermined DNA or the like in a solution. The number of specifically to be detected DNA sequences or the like depends on the number of working electrodes used.
BezugszeichenlisteLIST OF REFERENCE NUMBERS
1 Behälter1 container
OP1, 2, 3 erster, zweiter, dritter Operationsverstärker P PotentiostatOP1, 2, 3 first, second, third operational amplifier P potentiostat
Sl, 2, 3 erster, zweiter, dritter Strom-SpannungskonverterSl, 2, 3 first, second, third current-voltage converter
Rl, 2, 3 erster, zweiter, dritter WiderstandRl, 2, 3 first, second, third resistor
AD Messvorrichtung AEl, 2, 3 ArbeitselektrodenAD measuring device AEl, 2, 3 working electrodes
GE GegenelektrodeGE counter electrode
RE ReferenzelektrodeRE reference electrode
U Ausgang einer programmierbaren Spannungsquelle U Output of a programmable voltage source

Claims

Patentansprüche claims
1. Vorrichtung zum elektrochemischen Nachweis zumindest eines in einer Flüssigkeit enthaltenen biochemischen Moleküls aus einer Gruppe vorgegebener biochemischer Moleküle mit1. Device for the electrochemical detection of at least one biochemical molecule contained in a liquid from a group of predetermined biochemical molecules
einem mindestens eine Referenz- (RE) und mindestens eine Gegenelektrode (GE) sowie eine Vielzahl an Arbeitselektroden (AEl, AE2 , AE3) aufweisenden Mittel (1) zur Aufnahme der Flüssigkeit, wobei zum Nachweis jedes biochemischen Moleküls zumindest eine Arbeitselektrode (AEl, AE2 , AE3) vorgesehen ist, die mit einem zum jeweiligen biochemischen Molekül komplementären Molekül beschichtet ist, so dass die biochemischen Moleküle simultan nachweisbar sind,at least one reference (RE) and at least one counter electrode (GE) and a plurality of working electrodes (AE1, AE2, AE3) having means (1) for taking up the liquid, with at least one working electrode (AE1, AE2.) for the detection of each biochemical molecule , AE3) is provided, which is coated with a molecule complementary to the respective biochemical molecule, so that the biochemical molecules can be detected simultaneously,
einem Potentiostaten (P) zur Erzeugung eines vorgegebenen Spannungsverlaufs zwischen den Arbeitselektroden (AEl, AE2 , AE3) und der Referenzelektrode (RE) ,a potentiostat (P) for generating a predetermined voltage curve between the working electrodes (AE1, AE2, AE3) and the reference electrode (RE),
wobei jeder der Arbeitselektroden (AEl, AE2 , AE3) ein Strom-each of the working electrodes (AE1, AE2, AE3) a current
Spannungskonverter (Sl, S2, S3) nachgeschaltet ist, wobei die Strom-Spannungskonverter (Sl, S2 , S3) sämtliche Arbeitselektroden (AEl, AE2 , AE3) auf demselben Potenzial halten, undVoltage converter (Sl, S2, S3) is connected downstream, the current-voltage converter (Sl, S2, S3) keeping all working electrodes (AEl, AE2, AE3) at the same potential, and
einem Mittel (AD) zum Messen der durch die Arbeitselektroden (AEl, AE2, AE3) fließenden Ströme.a means (AD) for measuring the currents flowing through the working electrodes (AE1, AE2, AE3).
2. Vorrichtung nach Anspruch 1, wobei mehrere miteinander verbundene oder kapazitiv gekoppelte Referenzelektroden (RE) vorgesehen sind.2. Device according to claim 1, wherein a plurality of interconnected or capacitively coupled reference electrodes (RE) are provided.
3. Vorrichtung nach Anspruch 1 oder 2, wobei mehrere miteinander verbundene Gegenelektroden (GE) vorgesehen sind. 3. Device according to claim 1 or 2, wherein a plurality of interconnected counter electrodes (GE) are provided.
4. Vorrichtung nach einem der vorhergehenden Ansprüche, wobei das Mittel (AD) zum Messen einen Analog-Digital-Wandler aufweist .4. Device according to one of the preceding claims, wherein the means (AD) for measuring comprises an analog-to-digital converter.
5. Vorrichtung nach einem der vorhergehenden Ansprüche, wobei der Strom-Spannungskonverter (Sl, S2 , S3) ein einen ersten Operationsverstärker (OP1) aufweisenden Stromfolger ist, wobei ein nichtinvertierender Eingang (OP1+) des ersten Operationsverstärkers (OP1) an Masse anliegt und dessen inver- tierender Eingang (OP1-) über einen ersten Widerstand (Rl) mit dem Ausgang des ersten Operationsverstärkers (OP1) und mit der Arbeitselektrode (AEl) verbunden ist.5. Device according to one of the preceding claims, wherein the current-voltage converter (S1, S2, S3) is a current follower having a first operational amplifier (OP1), a non-inverting input (OP1 +) of the first operational amplifier (OP1) being connected to ground and the latter inverting input (OP1-) is connected via a first resistor (Rl) to the output of the first operational amplifier (OP1) and to the working electrode (AEl).
6. Vorrichtung nach einem der vorhergehenden Ansprüche, wo- bei parallel zum ersten Widerstand (Rl) eine Kapazität geschaltet ist.6. Device according to one of the preceding claims, wherein a capacitance is connected in parallel with the first resistor (R1).
7. Vorrichtung nach einem der vorhergehenden Ansprüche, wobei zur Einstellung des Strommessbereichs unterschiedlich große erste Widerstände (Rl) zwischen den invertierenden Eingang (OP1-) und den Ausgang des ersten Operationsverstärkers (0P1) einschaltbar sind.7. Device according to one of the preceding claims, wherein for setting the current measuring range different sized first resistors (R1) between the inverting input (OP1-) and the output of the first operational amplifier (0P1) can be switched on.
8. Vorrichtung nach einem der vorhergehenden Ansprüche, wo- bei das nachzuweisende biochemische Molekül eine Nukleinsäure und die komplementären biochemischen Moleküle zur nachzuweisenden Nukleinsäure komplementäre Nukleinsäuren sind.8. Device according to one of the preceding claims, wherein the biochemical molecule to be detected is a nucleic acid and the complementary biochemical molecules are nucleic acids complementary to the nucleic acid to be detected.
9. Vorrichtung nach einem der vorhergehenden Ansprüche, wo- bei der Potentiostat (P) einen als Spannungsfolger geschalteten zweiten Operationsverstärker (OP2) aufweist, an dessen nichtinvertierendem Eingang (OP2+) die Referenzelektrode (RE) angeschlossen ist . 9. Device according to one of the preceding claims, wherein the potentiostat (P) has a second operational amplifier (OP2) connected as a voltage follower, to whose non-inverting input (OP2 +) the reference electrode (RE) is connected.
10. Vorrichtung nach einem der vorhergehenden Ansprüche, wobei der Potentiostat (P) einen dritten Operationsverstärker (OP3) aufweist, an dessen Ausgang die Gegenelektrode (GE) angeschlossen ist, dessen invertierender Eingang (OP3-) über einen zweiten Widerstand (R2) mit dem Ausgang des zweiten10. Device according to one of the preceding claims, wherein the potentiostat (P) has a third operational amplifier (OP3), to the output of which the counter electrode (GE) is connected, the inverting input (OP3-) via a second resistor (R2) with the Exit of the second
Operationsverstärkers (OP2) verbunden und über einen dritten Widerstand (R3) an einer Einrichtung zur Erzeugung einer wählbaren Sollspannung angeschlossen ist, und wobei der nichtinvertierende Eingang (OP3+) des dritten Operationsver- stärkers (OP3) an Masse anliegt.Operational amplifier (OP2) connected and connected via a third resistor (R3) to a device for generating a selectable target voltage, and wherein the non-inverting input (OP3 +) of the third operational amplifier (OP3) is connected to ground.
11. Vorrichtung nach einem der vorhergehenden Ansprüche, wobei zwischen dem Ausgang des dritten Operationsverstärkers11. Device according to one of the preceding claims, wherein between the output of the third operational amplifier
(OP3) und dessen invertierenden Eingang (0P3-) eine Kapazität eingeschaltet ist.(OP3) and its inverting input (0P3-) a capacitance is switched on.
12. Verfahren zum elektrochemischen Nachweis zumindest eines in einer Flüssigkeit enthaltenen biochemischen Moleküls aus einer Gruppe vorgegebener biochemischer Moleküle mit folgen- den Schritten:12. Method for the electrochemical detection of at least one biochemical molecule contained in a liquid from a group of predetermined biochemical molecules with the following steps:
a) Bereitstellen eines Mittels (1) zur Aufnahme der Flüssigkeit, wobei das Mittel (1) mindestens eine Gegen- (GE) und eine Referenzelektrode (RE) sowie eine Vielzahl von Arbeitse- lektroden (AEl, AE2 , AE3) aufweist, wobei zum Nachweis jedes biochemischen Moleküls zumindest eine Arbeitselektrode (AEl, AE2 , AE3) vorgesehen ist, die mit einem zum jeweiligen biochemischen Molekül komplementären Molekül beschichtet ist, so dass die biochemischen Moleküle simultan nachweisbar sind,a) Providing a means (1) for receiving the liquid, the means (1) having at least one counter (GE) and one reference electrode (RE) and a plurality of working electrodes (AE1, AE2, AE3), with the Detection of each biochemical molecule, at least one working electrode (AE1, AE2, AE3) is provided, which is coated with a molecule that is complementary to the respective biochemical molecule, so that the biochemical molecules can be detected simultaneously,
b) Inkontaktbringen der Flüssigkeit mit den Arbeits- (AEl, AE2, AE3) , Gegen- (GE) und Referenzelektroden (RE) , c) gleichzeitiges Anlegen eines vorgegebenen Spannungsverlaufs zwischen den Arbeitselektroden (AEl, AE2 , AE3) und der Referenzelektrode (RE) undb) bringing the liquid into contact with the working (AE1, AE2, AE3), counter (GE) and reference electrodes (RE), c) simultaneous application of a predetermined voltage curve between the working electrodes (AE1, AE2, AE3) and the reference electrode (RE) and
d) Messen der durch die Arbeitselektroden (AEl, AE2 , AE3) fließenden Ströme, wobei während der Messung sämtliche Arbeitselektroden (AEl, AE2 , AE3) auf demselben Potenzial gehalten werden.d) measuring the currents flowing through the working electrodes (AE1, AE2, AE3), all working electrodes (AE1, AE2, AE3) being kept at the same potential during the measurement.
13. Verfahren nach Anspruch 13, wobei das Messen parallel oder mittels Multiplexen durchgeführt wird.13. The method according to claim 13, wherein the measurement is carried out in parallel or by means of multiplexing.
14. Verfahren nach einem der Ansprüche 13 oder 14, wobei die zwischen den Arbeitselektroden (AEl, AE2 , AE3) und der Refe- renzelektrode (RE) anliegende Spannung mit einem Potentiostaten (P) geregelt wird. 14. The method according to any one of claims 13 or 14, wherein the voltage present between the working electrodes (AE1, AE2, AE3) and the reference electrode (RE) is regulated with a potentiostat (P).
EP03740302A 2002-06-28 2003-06-23 Electrochemical detection method and device Expired - Lifetime EP1518123B1 (en)

Applications Claiming Priority (5)

Application Number Priority Date Filing Date Title
DE10229210 2002-06-28
DE10229210A DE10229210A1 (en) 2002-06-28 2002-06-28 Device for the detection of an analyte
DE2002129374 DE10229374C1 (en) 2002-06-29 2002-06-29 Electrochemical unit detecting biochemical molecules in fluid, includes electrodes coated with complementary biochemical molecules
DE10229374 2002-06-29
PCT/EP2003/006566 WO2004003556A1 (en) 2002-06-28 2003-06-23 Electrochemical detection method and device

Publications (2)

Publication Number Publication Date
EP1518123A1 true EP1518123A1 (en) 2005-03-30
EP1518123B1 EP1518123B1 (en) 2008-10-29

Family

ID=30001494

Family Applications (1)

Application Number Title Priority Date Filing Date
EP03740302A Expired - Lifetime EP1518123B1 (en) 2002-06-28 2003-06-23 Electrochemical detection method and device

Country Status (7)

Country Link
US (1) US7169289B2 (en)
EP (1) EP1518123B1 (en)
JP (1) JP2005531759A (en)
AT (1) ATE412899T1 (en)
AU (1) AU2003279777A1 (en)
DE (1) DE50310708D1 (en)
WO (1) WO2004003556A1 (en)

Families Citing this family (157)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6862465B2 (en) 1997-03-04 2005-03-01 Dexcom, Inc. Device and method for determining analyte levels
US8527026B2 (en) 1997-03-04 2013-09-03 Dexcom, Inc. Device and method for determining analyte levels
US6001067A (en) 1997-03-04 1999-12-14 Shults; Mark C. Device and method for determining analyte levels
US7657297B2 (en) * 2004-05-03 2010-02-02 Dexcom, Inc. Implantable analyte sensor
US7192450B2 (en) 2003-05-21 2007-03-20 Dexcom, Inc. Porous membranes for use with implantable devices
US6036924A (en) 1997-12-04 2000-03-14 Hewlett-Packard Company Cassette of lancet cartridges for sampling blood
US6391005B1 (en) 1998-03-30 2002-05-21 Agilent Technologies, Inc. Apparatus and method for penetration with shaft having a sensor for sensing penetration depth
US6949816B2 (en) 2003-04-21 2005-09-27 Motorola, Inc. Semiconductor component having first surface area for electrically coupling to a semiconductor chip and second surface area for electrically coupling to a substrate, and method of manufacturing same
US6175752B1 (en) * 1998-04-30 2001-01-16 Therasense, Inc. Analyte monitoring device and methods of use
US8688188B2 (en) 1998-04-30 2014-04-01 Abbott Diabetes Care Inc. Analyte monitoring device and methods of use
US9066695B2 (en) 1998-04-30 2015-06-30 Abbott Diabetes Care Inc. Analyte monitoring device and methods of use
US8480580B2 (en) 1998-04-30 2013-07-09 Abbott Diabetes Care Inc. Analyte monitoring device and methods of use
US8346337B2 (en) 1998-04-30 2013-01-01 Abbott Diabetes Care Inc. Analyte monitoring device and methods of use
US8974386B2 (en) 1998-04-30 2015-03-10 Abbott Diabetes Care Inc. Analyte monitoring device and methods of use
US8465425B2 (en) 1998-04-30 2013-06-18 Abbott Diabetes Care Inc. Analyte monitoring device and methods of use
US8641644B2 (en) 2000-11-21 2014-02-04 Sanofi-Aventis Deutschland Gmbh Blood testing apparatus having a rotatable cartridge with multiple lancing elements and testing means
DE10057832C1 (en) 2000-11-21 2002-02-21 Hartmann Paul Ag Blood analysis device has syringe mounted in casing, annular mounting carrying needles mounted behind test strip and being swiveled so that needle can be pushed through strip and aperture in casing to take blood sample
US6560471B1 (en) 2001-01-02 2003-05-06 Therasense, Inc. Analyte monitoring device and methods of use
EP1404234B1 (en) 2001-06-12 2011-02-09 Pelikan Technologies Inc. Apparatus for improving success rate of blood yield from a fingerstick
US9427532B2 (en) 2001-06-12 2016-08-30 Sanofi-Aventis Deutschland Gmbh Tissue penetration device
US7981056B2 (en) 2002-04-19 2011-07-19 Pelikan Technologies, Inc. Methods and apparatus for lancet actuation
AU2002315177A1 (en) 2001-06-12 2002-12-23 Pelikan Technologies, Inc. Self optimizing lancing device with adaptation means to temporal variations in cutaneous properties
US9226699B2 (en) 2002-04-19 2016-01-05 Sanofi-Aventis Deutschland Gmbh Body fluid sampling module with a continuous compression tissue interface surface
WO2002100460A2 (en) 2001-06-12 2002-12-19 Pelikan Technologies, Inc. Electric lancet actuator
US7041068B2 (en) 2001-06-12 2006-05-09 Pelikan Technologies, Inc. Sampling module device and method
ATE494837T1 (en) 2001-06-12 2011-01-15 Pelikan Technologies Inc INTEGRATED SYSTEM FOR BLOOD SAMPLE ANALYSIS WITH MULTIPLE-USE SAMPLING MODULE
US8337419B2 (en) 2002-04-19 2012-12-25 Sanofi-Aventis Deutschland Gmbh Tissue penetration device
US9795747B2 (en) 2010-06-02 2017-10-24 Sanofi-Aventis Deutschland Gmbh Methods and apparatus for lancet actuation
EP1404232B1 (en) 2001-06-12 2009-12-02 Pelikan Technologies Inc. Blood sampling apparatus and method
AU2002348683A1 (en) 2001-06-12 2002-12-23 Pelikan Technologies, Inc. Method and apparatus for lancet launching device integrated onto a blood-sampling cartridge
US6702857B2 (en) 2001-07-27 2004-03-09 Dexcom, Inc. Membrane for use with implantable devices
US20030032874A1 (en) 2001-07-27 2003-02-13 Dexcom, Inc. Sensor head for use with implantable devices
US7344894B2 (en) 2001-10-16 2008-03-18 Agilent Technologies, Inc. Thermal regulation of fluidic samples within a diagnostic cartridge
US8260393B2 (en) 2003-07-25 2012-09-04 Dexcom, Inc. Systems and methods for replacing signal data artifacts in a glucose sensor data stream
US8010174B2 (en) 2003-08-22 2011-08-30 Dexcom, Inc. Systems and methods for replacing signal artifacts in a glucose sensor data stream
US8858434B2 (en) 2004-07-13 2014-10-14 Dexcom, Inc. Transcutaneous analyte sensor
US9247901B2 (en) 2003-08-22 2016-02-02 Dexcom, Inc. Systems and methods for replacing signal artifacts in a glucose sensor data stream
US9282925B2 (en) 2002-02-12 2016-03-15 Dexcom, Inc. Systems and methods for replacing signal artifacts in a glucose sensor data stream
US7297122B2 (en) * 2002-04-19 2007-11-20 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US9314194B2 (en) 2002-04-19 2016-04-19 Sanofi-Aventis Deutschland Gmbh Tissue penetration device
US7547287B2 (en) 2002-04-19 2009-06-16 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US8579831B2 (en) 2002-04-19 2013-11-12 Sanofi-Aventis Deutschland Gmbh Method and apparatus for penetrating tissue
US7331931B2 (en) 2002-04-19 2008-02-19 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7481776B2 (en) 2002-04-19 2009-01-27 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7410468B2 (en) 2002-04-19 2008-08-12 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7291117B2 (en) 2002-04-19 2007-11-06 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7563232B2 (en) 2002-04-19 2009-07-21 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7976476B2 (en) 2002-04-19 2011-07-12 Pelikan Technologies, Inc. Device and method for variable speed lancet
US9248267B2 (en) 2002-04-19 2016-02-02 Sanofi-Aventis Deustchland Gmbh Tissue penetration device
US7901362B2 (en) 2002-04-19 2011-03-08 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7141058B2 (en) 2002-04-19 2006-11-28 Pelikan Technologies, Inc. Method and apparatus for a body fluid sampling device using illumination
US7582099B2 (en) 2002-04-19 2009-09-01 Pelikan Technologies, Inc Method and apparatus for penetrating tissue
US7491178B2 (en) 2002-04-19 2009-02-17 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7232451B2 (en) 2002-04-19 2007-06-19 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7648468B2 (en) 2002-04-19 2010-01-19 Pelikon Technologies, Inc. Method and apparatus for penetrating tissue
US7892183B2 (en) 2002-04-19 2011-02-22 Pelikan Technologies, Inc. Method and apparatus for body fluid sampling and analyte sensing
US7674232B2 (en) 2002-04-19 2010-03-09 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US8360992B2 (en) 2002-04-19 2013-01-29 Sanofi-Aventis Deutschland Gmbh Method and apparatus for penetrating tissue
US7708701B2 (en) 2002-04-19 2010-05-04 Pelikan Technologies, Inc. Method and apparatus for a multi-use body fluid sampling device
US7229458B2 (en) 2002-04-19 2007-06-12 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US8372016B2 (en) * 2002-04-19 2013-02-12 Sanofi-Aventis Deutschland Gmbh Method and apparatus for body fluid sampling and analyte sensing
US7717863B2 (en) 2002-04-19 2010-05-18 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US8267870B2 (en) 2002-04-19 2012-09-18 Sanofi-Aventis Deutschland Gmbh Method and apparatus for body fluid sampling with hybrid actuation
US7909778B2 (en) 2002-04-19 2011-03-22 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7371247B2 (en) 2002-04-19 2008-05-13 Pelikan Technologies, Inc Method and apparatus for penetrating tissue
US8702624B2 (en) 2006-09-29 2014-04-22 Sanofi-Aventis Deutschland Gmbh Analyte measurement device with a single shot actuator
US8784335B2 (en) 2002-04-19 2014-07-22 Sanofi-Aventis Deutschland Gmbh Body fluid sampling device with a capacitive sensor
US7175642B2 (en) 2002-04-19 2007-02-13 Pelikan Technologies, Inc. Methods and apparatus for lancet actuation
US9795334B2 (en) 2002-04-19 2017-10-24 Sanofi-Aventis Deutschland Gmbh Method and apparatus for penetrating tissue
US7374544B2 (en) 2002-04-19 2008-05-20 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US7524293B2 (en) 2002-04-19 2009-04-28 Pelikan Technologies, Inc. Method and apparatus for penetrating tissue
US8221334B2 (en) 2002-04-19 2012-07-17 Sanofi-Aventis Deutschland Gmbh Method and apparatus for penetrating tissue
US8574895B2 (en) 2002-12-30 2013-11-05 Sanofi-Aventis Deutschland Gmbh Method and apparatus using optical techniques to measure analyte levels
US7134999B2 (en) * 2003-04-04 2006-11-14 Dexcom, Inc. Optimized sensor geometry for an implantable glucose sensor
WO2004103147A2 (en) * 2003-05-02 2004-12-02 Pelikan Technologies, Inc. Method and apparatus for a tissue penetrating device user interface
US7875293B2 (en) * 2003-05-21 2011-01-25 Dexcom, Inc. Biointerface membranes incorporating bioactive agents
ATE476137T1 (en) 2003-05-30 2010-08-15 Pelikan Technologies Inc METHOD AND DEVICE FOR INJECTING LIQUID
WO2004107964A2 (en) 2003-06-06 2004-12-16 Pelikan Technologies, Inc. Blood harvesting device with electronic control
WO2006001797A1 (en) 2004-06-14 2006-01-05 Pelikan Technologies, Inc. Low pain penetrating
EP1635700B1 (en) 2003-06-13 2016-03-09 Sanofi-Aventis Deutschland GmbH Apparatus for a point of care device
JP2007500336A (en) 2003-07-25 2007-01-11 デックスコム・インコーポレーテッド Electrode system for electrochemical sensors
US8423113B2 (en) 2003-07-25 2013-04-16 Dexcom, Inc. Systems and methods for processing sensor data
US7519408B2 (en) 2003-11-19 2009-04-14 Dexcom, Inc. Integrated receiver for continuous analyte sensor
US7955261B2 (en) 2003-08-01 2011-06-07 Dexcom, Inc. System and methods for processing analyte sensor data
US8160669B2 (en) 2003-08-01 2012-04-17 Dexcom, Inc. Transcutaneous analyte sensor
US6931327B2 (en) 2003-08-01 2005-08-16 Dexcom, Inc. System and methods for processing analyte sensor data
US8369919B2 (en) 2003-08-01 2013-02-05 Dexcom, Inc. Systems and methods for processing sensor data
US9135402B2 (en) 2007-12-17 2015-09-15 Dexcom, Inc. Systems and methods for processing sensor data
US7494465B2 (en) 2004-07-13 2009-02-24 Dexcom, Inc. Transcutaneous analyte sensor
US20070208245A1 (en) * 2003-08-01 2007-09-06 Brauker James H Transcutaneous analyte sensor
US7591801B2 (en) 2004-02-26 2009-09-22 Dexcom, Inc. Integrated delivery device for continuous glucose sensor
US8622905B2 (en) 2003-08-01 2014-01-07 Dexcom, Inc. System and methods for processing analyte sensor data
US7774145B2 (en) 2003-08-01 2010-08-10 Dexcom, Inc. Transcutaneous analyte sensor
US8845536B2 (en) 2003-08-01 2014-09-30 Dexcom, Inc. Transcutaneous analyte sensor
US20190357827A1 (en) 2003-08-01 2019-11-28 Dexcom, Inc. Analyte sensor
US8761856B2 (en) 2003-08-01 2014-06-24 Dexcom, Inc. System and methods for processing analyte sensor data
US8275437B2 (en) 2003-08-01 2012-09-25 Dexcom, Inc. Transcutaneous analyte sensor
US20140121989A1 (en) 2003-08-22 2014-05-01 Dexcom, Inc. Systems and methods for processing analyte sensor data
US7920906B2 (en) 2005-03-10 2011-04-05 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US8282576B2 (en) 2003-09-29 2012-10-09 Sanofi-Aventis Deutschland Gmbh Method and apparatus for an improved sample capture device
WO2005037095A1 (en) 2003-10-14 2005-04-28 Pelikan Technologies, Inc. Method and apparatus for a variable user interface
US20050090607A1 (en) * 2003-10-28 2005-04-28 Dexcom, Inc. Silicone composition for biocompatible membrane
US9247900B2 (en) 2004-07-13 2016-02-02 Dexcom, Inc. Analyte sensor
US8774886B2 (en) 2006-10-04 2014-07-08 Dexcom, Inc. Analyte sensor
US8287453B2 (en) 2003-12-05 2012-10-16 Dexcom, Inc. Analyte sensor
US11633133B2 (en) 2003-12-05 2023-04-25 Dexcom, Inc. Dual electrode system for a continuous analyte sensor
US8423114B2 (en) 2006-10-04 2013-04-16 Dexcom, Inc. Dual electrode system for a continuous analyte sensor
ATE480761T1 (en) * 2003-12-05 2010-09-15 Dexcom Inc CALIBRATION METHODS FOR A CONTINUOUSLY WORKING ANALYTICAL SENSOR
US8364231B2 (en) 2006-10-04 2013-01-29 Dexcom, Inc. Analyte sensor
EP3263032B1 (en) 2003-12-09 2024-01-24 Dexcom, Inc. Signal processing for continuous analyte sensor
US7822454B1 (en) 2005-01-03 2010-10-26 Pelikan Technologies, Inc. Fluid sampling device with improved analyte detecting member configuration
US8668656B2 (en) 2003-12-31 2014-03-11 Sanofi-Aventis Deutschland Gmbh Method and apparatus for improving fluidic flow and sample capture
US7637868B2 (en) * 2004-01-12 2009-12-29 Dexcom, Inc. Composite material for implantable device
US8808228B2 (en) 2004-02-26 2014-08-19 Dexcom, Inc. Integrated medicament delivery device for use with continuous analyte sensor
US20050245799A1 (en) * 2004-05-03 2005-11-03 Dexcom, Inc. Implantable analyte sensor
US8792955B2 (en) 2004-05-03 2014-07-29 Dexcom, Inc. Transcutaneous analyte sensor
US8277713B2 (en) * 2004-05-03 2012-10-02 Dexcom, Inc. Implantable analyte sensor
WO2006011062A2 (en) 2004-05-20 2006-02-02 Albatros Technologies Gmbh & Co. Kg Printable hydrogel for biosensors
US9820684B2 (en) 2004-06-03 2017-11-21 Sanofi-Aventis Deutschland Gmbh Method and apparatus for a fluid sampling device
US9775553B2 (en) 2004-06-03 2017-10-03 Sanofi-Aventis Deutschland Gmbh Method and apparatus for a fluid sampling device
US7640048B2 (en) 2004-07-13 2009-12-29 Dexcom, Inc. Analyte sensor
US20080242961A1 (en) * 2004-07-13 2008-10-02 Dexcom, Inc. Transcutaneous analyte sensor
US8452368B2 (en) 2004-07-13 2013-05-28 Dexcom, Inc. Transcutaneous analyte sensor
US20070045902A1 (en) 2004-07-13 2007-03-01 Brauker James H Analyte sensor
US7783333B2 (en) * 2004-07-13 2010-08-24 Dexcom, Inc. Transcutaneous medical device with variable stiffness
US8565848B2 (en) 2004-07-13 2013-10-22 Dexcom, Inc. Transcutaneous analyte sensor
JP4742544B2 (en) * 2004-09-08 2011-08-10 凸版印刷株式会社 Gene check apparatus and method
US20060167382A1 (en) * 2004-12-30 2006-07-27 Ajay Deshmukh Method and apparatus for storing an analyte sampling and measurement device
US8652831B2 (en) 2004-12-30 2014-02-18 Sanofi-Aventis Deutschland Gmbh Method and apparatus for analyte measurement test time
US20090054811A1 (en) * 2004-12-30 2009-02-26 Dirk Boecker Method and apparatus for analyte measurement test time
EP1842052A1 (en) * 2005-01-26 2007-10-10 Rapid Laboratory Microsystems Inc. Method, system and device for obtaining electrochemical measurements
CN102385260B (en) * 2005-02-10 2014-11-05 Asml荷兰有限公司 Immersion liquid, exposure apparatus, and exposure process
US8060174B2 (en) 2005-04-15 2011-11-15 Dexcom, Inc. Analyte sensing biointerface
EP1991110B1 (en) 2006-03-09 2018-11-07 DexCom, Inc. Systems and methods for processing analyte sensor data
US20080071157A1 (en) 2006-06-07 2008-03-20 Abbott Diabetes Care, Inc. Analyte monitoring system and method
US8562558B2 (en) 2007-06-08 2013-10-22 Dexcom, Inc. Integrated medicament delivery device for use with continuous analyte sensor
EP4098177A1 (en) 2007-10-09 2022-12-07 DexCom, Inc. Integrated insulin delivery system with continuous glucose sensor
US8417312B2 (en) 2007-10-25 2013-04-09 Dexcom, Inc. Systems and methods for processing sensor data
FR2923296B1 (en) * 2007-11-02 2009-11-20 Commissariat Energie Atomique MULTI-CHANNEL POTENTIOSTAT HAVING AN ADJUSTABLE COUNTER-ELECTRODE POTENTIAL.
US9839395B2 (en) 2007-12-17 2017-12-12 Dexcom, Inc. Systems and methods for processing sensor data
US20090209883A1 (en) * 2008-01-17 2009-08-20 Michael Higgins Tissue penetrating apparatus
US9143569B2 (en) 2008-02-21 2015-09-22 Dexcom, Inc. Systems and methods for processing, transmitting and displaying sensor data
US8396528B2 (en) 2008-03-25 2013-03-12 Dexcom, Inc. Analyte sensor
WO2009126900A1 (en) 2008-04-11 2009-10-15 Pelikan Technologies, Inc. Method and apparatus for analyte detecting device
US9375169B2 (en) 2009-01-30 2016-06-28 Sanofi-Aventis Deutschland Gmbh Cam drive for managing disposable penetrating member actions with a single motor and motor and control system
EP2410910A4 (en) 2009-03-27 2014-10-15 Dexcom Inc Methods and systems for promoting glucose management
US20110027453A1 (en) 2009-07-02 2011-02-03 Dexcom, Inc. Continuous analyte sensors and methods of making same
WO2011047271A1 (en) * 2009-10-16 2011-04-21 Microchips, Inc. Multi-channel potentiostat for biosensor arrays
US8965476B2 (en) 2010-04-16 2015-02-24 Sanofi-Aventis Deutschland Gmbh Tissue penetration device
EP2697650B1 (en) 2011-04-15 2020-09-30 Dexcom, Inc. Advanced analyte sensor calibration and error detection
JP5689020B2 (en) * 2011-05-12 2015-03-25 日置電機株式会社 Trace component detection apparatus and trace component detection method
CZ2014672A3 (en) * 2014-09-30 2015-08-12 Vysoké Učení Technické V Brně Potentiostat
US9983167B2 (en) * 2014-10-31 2018-05-29 Zansors, Llc Multichannel potentiostat analyzer system and methods
JP7231963B2 (en) * 2017-02-21 2023-03-02 国立大学法人山形大学 Electrochemical measuring device
WO2018225058A1 (en) * 2017-06-04 2018-12-13 B.G. Negev Technologies And Applications Ltd., At Ben-Gurion University Electrochemical detection device and method
US11331022B2 (en) 2017-10-24 2022-05-17 Dexcom, Inc. Pre-connected analyte sensors
CN212438615U (en) 2017-10-24 2021-02-02 德克斯康公司 Wearable device

Family Cites Families (27)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US792231A (en) * 1904-04-01 1905-06-13 Nat Tube Co Art of cross-rolling tubular bodies or blanks in a heated state.
US2176155A (en) * 1936-09-03 1939-10-17 Timken Roller Bearing Co Apparatus for elongating and decreasing the wall thickness of tubular blanks
FR2266557B1 (en) * 1974-04-05 1977-10-14 Valti Fabr Tubes Roulements
DE2652759C3 (en) * 1976-11-16 1980-05-22 Mannesmann Ag, 4000 Duesseldorf Detachable connection between a mandrel and the mandrel bar of a cross rolling mill
JPS5913924B2 (en) * 1979-12-25 1984-04-02 日本鋼管株式会社 Core metal for piercing rolling mill
US4315753A (en) * 1980-08-14 1982-02-16 The United States Of America As Represented By The Secretary Of The Interior Electrochemical apparatus for simultaneously monitoring two gases
DE3114177C2 (en) * 1981-04-03 1984-08-23 Mannesmann AG, 4000 Düsseldorf Process for the production of a working tool for non-cutting hot forming of steel and hot working tool
US4488556A (en) * 1982-06-03 1984-12-18 Critikon, Inc. AC Mode operation of chemfet devices
US4655880A (en) * 1983-08-01 1987-04-07 Case Western Reserve University Apparatus and method for sensing species, substances and substrates using oxidase
JPS63101744A (en) * 1986-10-18 1988-05-06 Matsushita Electric Works Ltd Assay apparatus for component in liquid
IL82131A0 (en) * 1987-04-07 1987-10-30 Univ Ramot Coulometric assay system
JPH02224806A (en) * 1989-02-28 1990-09-06 Nkk Corp Plug for seamless steel pipe manufacturing
JPH04264246A (en) * 1991-02-19 1992-09-21 Matsushita Electric Ind Co Ltd biosensor
US5217112A (en) * 1991-09-04 1993-06-08 Almon Amy C Voltametric analysis apparatus and method
DE4136779A1 (en) 1991-11-08 1993-05-13 Bayer Ag DEVICE FOR SIMULTANEOUS DETECTION OF DIFFERENT GAS COMPONENTS
US5260663A (en) * 1992-07-14 1993-11-09 Anatel Corporation Methods and circuits for measuring the conductivity of solutions
DE4424355C2 (en) 1994-07-11 1996-07-18 Fraunhofer Ges Forschung Electrochemical analysis method
JPH10180315A (en) * 1996-12-27 1998-07-07 Kawasaki Steel Corp Plug for seamless steel pipe rolling and method for manufacturing seamless steel pipe
JP3271547B2 (en) * 1997-01-23 2002-04-02 ダイキン工業株式会社 Sensor device
US7297312B2 (en) * 1998-03-16 2007-11-20 University Of Cincinnati Simultaneous multianalyte electrochemical assay based on spatial resolution
JP3214561B2 (en) * 1998-07-02 2001-10-02 日本電気株式会社 Enzyme electrode, biosensor and measuring device using the same
JP2001021526A (en) * 1999-07-02 2001-01-26 Akebono Brake Res & Dev Center Ltd Measuring method of sample solution by using biosensor
DE10015818A1 (en) 2000-03-30 2001-10-18 Infineon Technologies Ag Biosensor and method for determining macromolecular biopolymers using a biosensor
JP2002017352A (en) * 2000-04-28 2002-01-22 Fuji Photo Film Co Ltd Tool for detecting nucleic acid specimen and method for electrochemical detection
US6818109B2 (en) * 2000-09-29 2004-11-16 Kabushiki Kaisha Toshiba Nucleic acid detections sensor
US20030022150A1 (en) * 2001-07-24 2003-01-30 Sampson Jeffrey R. Methods for detecting a target molecule
CN100404151C (en) * 2002-12-12 2008-07-23 住友金属工业株式会社 Method for manufacturing seamless metal pipe

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO2004003556A1 *

Also Published As

Publication number Publication date
DE50310708D1 (en) 2008-12-11
WO2004003556A1 (en) 2004-01-08
US7169289B2 (en) 2007-01-30
ATE412899T1 (en) 2008-11-15
EP1518123B1 (en) 2008-10-29
JP2005531759A (en) 2005-10-20
US20050211571A1 (en) 2005-09-29
AU2003279777A1 (en) 2004-01-19

Similar Documents

Publication Publication Date Title
EP1518123B1 (en) Electrochemical detection method and device
EP0527126B1 (en) Electrochemical determination of oxygen concentration
DE69623006T2 (en) SENSOR
DE60107224T2 (en) ELECTROCHEMICAL BIOSENSOR CALIBRATION DEVICE
EP1516175A1 (en) Biosensor array and method for operating a biosensor array
DE112011105207T5 (en) Biomolecule information analysis apparatus
EP1789811B1 (en) Biosensor array and method for detecting a sensor event
DE10324912A1 (en) Method for the detection of DNA point mutations (SNP analysis) and associated arrangement
DE69422856T2 (en) Circuit arrangement and method for determining the current through a sensor
DE3033730C2 (en) Device for detecting chemical substances
DE102004031371A1 (en) Monolithically integrated sensor arrangement, sensor array and method for producing a monolithically integrated sensor arrangement
EP0717841A1 (en) Electrochemical analysis process
DE10224567B4 (en) Sensor arrangement and method for operating a sensor arrangement
EP1328799B1 (en) Electronic circuit, sensor arrangement and method for processing a sensor signal
DE10229374C1 (en) Electrochemical unit detecting biochemical molecules in fluid, includes electrodes coated with complementary biochemical molecules
EP0060533B1 (en) Electrochemical analyser
DE102004031370B4 (en) Apparatus and method for emulating a counter electrode in a monolithic integrated electrochemical analysis system
Komorsky‐Lovrić et al. A peak current–scan rate relationship in staircase voltammetry of a surface redox reaction
DE102006006347B3 (en) Sensor device for an electrochemical measuring device and method for carrying out electrochemical measurements
EP1208239B1 (en) Method for detecting and quantifying first biopolymers that are located in a liquid
EP2264445B1 (en) Coulometric humidity sensor and corresponding method
AT407199B (en) pH sensor
DE4424179C2 (en) UV-polymerizable enzyme paste for the production of thick-film biosensors and thick-film biosensors produced therewith
DE19739317A1 (en) Electrical circuit for an electrochemical sensor
DE2633941A1 (en) ANALYZER AND METHOD OF ANALYSIS

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

17P Request for examination filed

Effective date: 20041129

AK Designated contracting states

Kind code of ref document: A1

Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HU IE IT LI LU MC NL PT RO SE SI SK TR

AX Request for extension of the european patent

Extension state: AL LT LV MK

DAX Request for extension of the european patent (deleted)
GRAP Despatch of communication of intention to grant a patent

Free format text: ORIGINAL CODE: EPIDOSNIGR1

GRAS Grant fee paid

Free format text: ORIGINAL CODE: EPIDOSNIGR3

GRAA (expected) grant

Free format text: ORIGINAL CODE: 0009210

AK Designated contracting states

Kind code of ref document: B1

Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HU IE IT LI LU MC NL PT RO SE SI SK TR

REG Reference to a national code

Ref country code: GB

Ref legal event code: FG4D

Free format text: NOT ENGLISH

REG Reference to a national code

Ref country code: CH

Ref legal event code: EP

REG Reference to a national code

Ref country code: IE

Ref legal event code: FG4D

Free format text: LANGUAGE OF EP DOCUMENT: GERMAN

REF Corresponds to:

Ref document number: 50310708

Country of ref document: DE

Date of ref document: 20081211

Kind code of ref document: P

NLV1 Nl: lapsed or annulled due to failure to fulfill the requirements of art. 29p and 29m of the patents act
PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: ES

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20090209

Ref country code: BG

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20090129

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: PT

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20090330

Ref country code: FI

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

Ref country code: SI

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

Ref country code: NL

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

REG Reference to a national code

Ref country code: IE

Ref legal event code: FD4D

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: RO

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

Ref country code: IE

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

Ref country code: DK

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

Ref country code: EE

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: IT

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

Ref country code: SE

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20090129

Ref country code: CZ

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

PLBE No opposition filed within time limit

Free format text: ORIGINAL CODE: 0009261

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: NO OPPOSITION FILED WITHIN TIME LIMIT

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: SK

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

26N No opposition filed

Effective date: 20090730

BERE Be: lapsed

Owner name: NOVEMBER A.G. GESELLSCHAFT FUR MOLEKULARE MEDIZIN

Effective date: 20090630

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: MC

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20090630

REG Reference to a national code

Ref country code: CH

Ref legal event code: PL

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: LI

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20090630

Ref country code: CH

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20090630

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: BE

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20090630

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: AT

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20090623

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: GR

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20090130

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: LU

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20090623

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: HU

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20090430

PGFP Annual fee paid to national office [announced via postgrant information from national office to epo]

Ref country code: FR

Payment date: 20110621

Year of fee payment: 9

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: TR

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

PGFP Annual fee paid to national office [announced via postgrant information from national office to epo]

Ref country code: GB

Payment date: 20110622

Year of fee payment: 9

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: CY

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20081029

PGFP Annual fee paid to national office [announced via postgrant information from national office to epo]

Ref country code: DE

Payment date: 20110615

Year of fee payment: 9

GBPC Gb: european patent ceased through non-payment of renewal fee

Effective date: 20120623

REG Reference to a national code

Ref country code: FR

Ref legal event code: ST

Effective date: 20130228

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: FR

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20120702

Ref country code: DE

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20130101

Ref country code: GB

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20120623

REG Reference to a national code

Ref country code: DE

Ref legal event code: R119

Ref document number: 50310708

Country of ref document: DE

Effective date: 20130101